Funds are requested to purchase a Perkin Elmer Operetta benchtop automated high-throughput, high- content imaging system. At present, no such instrument is available to the Wistar Institute or the neighboring research community at the University of Pennsylvania or Drexel University. The Wistar Institute has recently established a Molecular Screening Facility to support screening of shRNA, siRNA, miRNA, and small molecule chemical libraries in microtiter plates. The facility has made great progress in developing assays that utilize whole-well readouts. However neither the Molecular Screening nor Microscopy Facility at the Wistar Institute have the capability to support high-content screening. Furthermore, a survey of the neighboring University of Pennsylvania and Drexel School of Medicine indicate that access to this technology as a shared resource to support screening experiments is inadequate in our research community. Numerous investigators at Wistar have requested the facility expand its capabilities into high- content cell-based screening assays. The requested Operetta will be located in the Wistar Institute Molecular Screening Facility, which was established in 2008 and is dedicated to maintaining automated, high-throughput technologies required to support systems biology approaches to identify and validate cellular targets and/or lead molecules to treat human diseases, including cancer, diabetes, inflammation, and atherosclerosis. The unique features of the Operetta will greatly expand imaging capabilities at Wistar to include automated high-throughput acquisition and quantitative analysis of image data. Most importantly the Operetta will immediately impact the ability of 8 Major NIH-funded users and a group of Minor users to develop cell-based high-throughput screening assays that focus on the analysis of cells and their molecular components. An internal advisory committee comprised of users and senior staff from the Wistar Institute will oversee the operation, maintenance, and equitable access to this instrument. In this application, we present projects and preliminary data from studies that require access to high-throughput, high-content imaging. These projects include development of miniaturized assays that utilize 3D organotypic cultures for high-throughput screening, assessment of cellular signaling networks that regulate wound healing and tissue regeneration, identification of cellular factors that regulate the architecture of chromatin domains and gene expression during latent Herpes Virus infection, and nucleocytoplasmic shuttling of cellular cofactors involved in epithelial-to mesenchymal transition, migration and tumor metastasis. The purchase of the Operetta will greatly expand the capabilities and versatility of the Wistar Molecular Screening Facility and Microscopy facilities to support transformative research aimed to reduce the burden of human disease.